Project Summary/ Abstract Neisseria gonorrhoeae causes both localized uncomplicated infections at mucosal surfaces and more invasive forms of disease including pelvic inflammatory disease and bloodstream infections. The ability of gonococci to evade the bactericidal action of human serum-antibody and complement (SAC) is important in invasive bloodstream infections, but the genetic and biochemical basis for this resistance is not fully understood. Our long-term goal is to define the molecular basis for SAC resistance expressed by gonococci so that novel strategies can be developed to prevent or treat bloodstream infections caused by Gram-negative pathogens. Gonococcal infections continue to be a major clinical problem for the veteran population, especially female veterans who often have severe gynecologic complications that result from invasive forms of disease. The results obtained from our investigations will help in understanding how gonococci and other Gram-negative pathogens can escape an important, innate host defense mechanism. A more complete understanding of how Gram-negative bacteria resist the bactericidal action of SAC is essential for the successful development of novel therapeutics or vaccines that would reduce the incidence and/or severity of bloodstream infections. We hypothesize that genes involved in lipooligosaccharide (LOS) biosynthesis and those that encode membrane proteins are important in determining the ability of gonococci to resist the bactericidal action of SAC. We have evidence that one mechanism by which gonococci stably resist killing by normal human serum (NHS) is genetically determined through structural alterations in the core oligosaccharide and lipid A components of the gonococcal LOS. We will now determine the repertoire of gonococcal genes that determine SAC resistance. We also have evidence that human serum triggers a response in SAC-resistant gonococci through a mechanism dependent on an active complement system. We will now define the gonococcal genes that are differentially expressed during bacterial growth in NHS and will determine the role of their gene products in determining SAC resistance. We will combine techniques of microbial genetics, molecular biology and biochemistry to identify and characterize the cell envelope components of gonococci that are important for SAC resistance. In Specific Aim 1, we will identify and characterize genes that determine SAC resistance in strains of gonococci isolated from the bloodstream of patients with disseminated gonococcal infection. We will also use transposon mutagenesis to identify genes in SAC-sensitive and -resistant strains that determine levels of bacterial susceptibility to this host defense system. In Specific Aim 2, we will use microarrays to identify genes in gonococci that are differentially expressed during exposure to and growth in human serum. Serum-regulated genes will be studied by constructing defined mutations in order to determine their role in SAC resistance. PUBLIC HEALTH RELEVANCE: Project Narrative - Relevance to Veterans Health Unfortunately, despite a downward trend in the total number of yearly cases of gonorrhea in the United States since the mid-1980s, the incidence of this infection remains at a distressingly high level in both the civilian and military populations. While the disease in all of its various forms can still be treated with antibiotics, the emergence of strains expressing resistance to inexpensive antibiotics (e.g., penicillin and tetracycline) has necessitated the use of more expensive drugs. The capacity of certain gonococcal strains to cause serious, invasive forms of disease increases health costs as a result of diagnostic and management expenses. With more female veterans using the Medical Service as their primary mode of care, it is likely that VA Medical Centers will experience the health care costs associated with invasive gonococcal infections.